Molecular imaging is frequently performed with contrast agents that have been specifically designed to target to cells or molecules associated with the disease. These agents also produce a distinct signal that can be detected by an imaging method. In this project, we develop a novel type of molecular contrast agent based on engineered protein microspheres functionalized to target the vascular endothelium in atherosclerotic cardiovascular disease. These agents are designed specifically for enhancing contrast in optical coherence tomography (OCT), an emerging high-resolution biomedical imaging modality that can be used for intravascular detection and assessment of atherosclerotic lesions. Objective/Hypothesis: Multifunctional microsphere contrast agents will be developed to provide molecularly-specific contrast enhancement in OCT. Our hypothesis is that these engineered microspheres, targeted to the 1v23 integrin, can provide molecular contrast in OCT for in vivo catheter-based localization of atherosclerotic lesions. Specific Aims: Aim 1: Optimize the optical scattering properties of multifunctional microspheres. Aim 2: Determine targeting efficiency of microspheres to the 1v23 integrin in in vitro cell culture. Aim 3: Characterize and quantify OCT imaging and molecular contrast enhancement in an ex vivo hyperlipidemic pre-clinical model of atherosclerosis using targeted microspheres. Aim 4: Characterize and quantify OCT imaging and molecular contrast enhancement in vivo in a hyperlipidemic pre-clinical model of atherosclerosis using targeted microspheres. Study Design: This project will involve interdisciplinary efforts including the chemistry of fabricating microsphere contrast agents, the engineering and optimization of their optical properties, the use of novel optical imaging technology and data acquisition systems, and immunological targeting concepts for applications in in vitro cell culture and ex vivo and in vivo pre-clinical models. Clinical Relevance: This research is highly significant in the areas of detection, imaging, and assessment of atherosclerotic cardiovascular disease. The novel use of high-resolution OCT for tracking the spatial localization of molecularly-targeted optical contrast agents in pre-clinical ex vivo experiments and in in vivo catheter-based imaging of atherosclerosis enables a new imaging methodology and paradigm in the detection, treatment, and monitoring of atherosclerotic cardiovascular disease. PUBLIC HEALTH RELEVANCE: This proposal describes the development and application of targeted multifunctional microspheres for enhancing contrast in optical coherence tomography (OCT), an emerging high-resolution biomedical imaging modality. Our hypothesis is that these engineered microspheres can provide molecularly-sensitive contrast for OCT. The use of these agents therefore offers the potential for improving the diagnostic utility of OCT for investigations in the detection, imaging, assessment, and monitoring of cardiovascular disease.